Characterisation of a hybrid histidine kinase controlling acetate metabolism in Acinetobacter baumannii

RNA sequencing transcriptomics was performed on a highly multidrug resistant A. baumannii strain belonging to international clone I, AB5075_UW and a transposon insertion inactivated mutant of ABUW_0182 (acmS), which encodes a hybrid histidine kinase.Transcriptomics suggests that AcmS controls expression of the genes involved in short-chain fatty acid metabolism in A. baumannii. Biophysical analyses showed ABUW_0182 binds acetic and propionic acid with affinities in a low micromolar range, suggesting they represent the physiological ligands for this hybrid histidine kinase system. Overall design: Global transcriptomic profiles of the acmS mutant and the parental A. baumannii strain were compared. The parental and mutant cells were grown to mid exponential phase at 37°C, with shaking. Cell pellets were harvested and the total RNA was extracted from three biological replicates. Ribosomal RNA was depleted using Ribozero magnetic kit (Bacteria) (Illumina, Inc., USA) and the cDNA library was generated using Truseq stranded RNA preparation kit (Illumina, Inc., USA). RNA libraries were prepared for sequencing using standard Illumina protocols.

本研究对一株属于国际克隆I的高度多重耐药鲍曼不动杆菌（A. baumannii）菌株AB5075_UW，以及其ABUW_0182（acmS）基因的转座子插入失活突变株开展了转录组RNA测序分析，其中acmS基因编码一种杂合组氨酸激酶。转录组分析结果显示，AcmS可调控鲍曼不动杆菌参与短链脂肪酸代谢的基因表达。生物物理分析表明，ABUW_0182蛋白可与乙酸和丙酸以低微摩尔级的亲和力结合，提示二者为该杂合组氨酸激酶系统的生理配体。实验整体设计：比较acmS突变株与亲本鲍曼不动杆菌菌株的全球转录组谱。将亲本菌株与突变株于37℃、振荡培养至指数生长中期，收集菌体沉淀，从三次生物学重复样本中提取总RNA。使用RiboZero细菌磁珠试剂盒（Illumina公司，美国）去除核糖体RNA，采用Truseq链特异性RNA建库试剂盒（Illumina公司，美国）构建cDNA文库。RNA文库按照标准Illumina流程制备并进行测序。